GDNF family ligand receptor components Ret and GFRalpha-1 in the human trigeminal ganglion and sensory nuclei

The occurrence of Ret and GFRalpha-1 receptors is shown by immunohistochemistry in the human trigeminal sensory system at pre-, postnatal and adult age. Receptor-labeled neurons occur in both trigeminal ganglion and mesencephalic nucleus. In adult trigeminal ganglion, about 75% of Ret- and 65% of GFRalpha-1-labeled neurons are small- and medium-sized. The proportion of Ret+ and GFRalpha-1+ trigeminal ganglion neurons in the adult is about 25 and 60%, respectively. The majority of Ret+ are double labeled for GFRalpha-1 and glial cell line-derived neurotrophic factor (GDNF). Most of the GFRalpha-1+ cells contain GDNF and about 50% of them contain Ret. Triple labeling shows many Ret+/GDNF+/GFRalpha-1+ neurons, but also a number of Ret-/GDNF+/GFRalpha-1+ and Ret+/GDNF-/GFRalpha-1+ cells. Both Ret+ and GFRalpha-1+ neuronal subpopulations overlap with that containing calcitonin gene-related peptide. Ret+ pericellular basket-like nerve fibers occur in the adult trigeminal ganglion. Centrally, immunoreactivity is restricted to the spinal nucleus pars caudalis and pars interpolaris and to the mesencephalic nucleus. In adult specimens, Ret+ nerve fibers and puncta gather in the inner substantia gelatinosa. Ret+ neurons occur in the spinal nucleus and are more frequent in newborn than in adult subjects. Central GFRalpha-1+-labeled neurons and punctate elements are sparse. These findings support the involvement of GDNF and possibly other cognate ligands in the trophism of human trigeminal primary sensory neurons from prenatal life to adulthood, indicating a selective commitment to cells devoted to protopathic and proprioceptive sensory transmission. They also support the possibility that receptor molecules other than Ret could be active in transducing the ligand signal.     

Glial cell line-derived neurotrophic factor increases calcitonin gene-related peptide immunoreactivity in sensory and motoneurons in vivo

Calcitonin gene-related peptide (CGRP) is expressed at high levels in roughly 50% of spinal sensory neurons and plays a role in peripheral vasodilation as well as nociceptive signalling in the spinal cord. Spinal motoneurons express low levels of CGRP; motoneuronal CGRP is thought to be involved in end-plate plasticity and to have trophic effects on target muscle cells. As both sensory and motoneurons express receptors for glial cell line-derived neurotrophic factor (GDNF) we sought to determine whether CGRP was regulated by GDNF. Rats were treated intrathecally for 1-3 weeks with recombinant human GDNF or nerve growth factor (NGF) (12 microg/day) and dorsal root ganglia and spinal cords were stained for CGRP. The GDNF treatment not only increased CGRP immunoreactivity in both sensory and motoneurons but also resulted in hypertrophy of both populations. By combined in situ hybridization and immunohistochemistry we found that, in the dorsal root ganglia, CGRP was up-regulated specifically in neurons expressing GDNF but not NGF receptors following GDNF treatment. Despite the increase in CGRP in GDNF-treated rats, there was no increase in thermal or mechanical pain sensitivity, while NGF-treated animals showed significant decreases in pain thresholds. In motoneurons, GDNF increased the overall intensity of CGRP immunoreactivity but did not increase the number of immunopositive cells. As GDNF has been shown to promote the regeneration of both sensory and motor axons, and as CGRP appears to be involved in motoneuronal plasticity, we reason that at least some of the regenerative effects of GDNF are mediated through CGRP up-regulation.     

Immunocytochemical localization of μ-opioid receptor in primary afferent neurons containing substance P or calcitonin gene-related peptide. A light and electron microscope study in the rat

Co-expression of μ-opioid receptor-like immunoreactivity (MOR-LI) with substance P (SP)- or calcitonin gene-related (CGRP)-LI was observed in rat trigeminal and dorsal root ganglion neurons. In particular, MOR-LI was found in axon terminals with SP- or CGRP-LI in laminae I and II of the medullary and spinal dorsal horns. MOR may be implicated in modulation of release of SP and CGRP from primary sensory afferents.     

Peripheral and central distribution of TRPV1, substance P and CGRP of rat corneal neurons

The rat corneal neurons expressing vanilloid receptor TRPV1, substance P (SP) and calcitonin-gene-related peptide (CGRP) were examined. In the cornea, some TRPV1-immunoreactive nerve fibers displayed either SP- or CGRP immunoreactivity also. For observing corneal neuronal elements in the trigeminal ganglion (TG) and in the medulla oblongata, retrograde and anterograde cholera toxin subunit B (CTB) tracing methods combining with triple immunofluorescence technique were performed. The corneal neuronal somata were located in the ophthalmic division of the TG; 37% of them were immunoreactive for TRPV1. One third and three quarters of the corneal TRPV1-immunoreactive neurons co-expressed SP and CGRP, respectively. All of SP-immunoreactive corneal neurons exhibited TRPV1 immunoreactivity. They were predominantly medium-sized (mean +/- SE = 638.2 +/- 49.5 microm(2)) and significantly larger than SP-immunoreactive and TRPV1-immunonegative neurons in the ophthalmic division of the TG. The central projection fibers of corneal neurons co-expressing TRPV1 with SP and CGRP were observed at the subnucleus interpolaris/caudalis transition within trigeminal nucleus. The present study suggests that TRPV1 of the corneal neurons works in close relation to SP and CGRP both in the cornea and CNS for healing and nociceptive transduction.     

The effect of streptozotocin-induced diabetes mellitus on substance P and calcitonin gene-related peptide expression in the rat trigeminal ganglion

Substance P (SP) and calcitonin gene-related peptide (CGRP) constitute the main sensory peptides in the trigeminal ganglion (TG). The objective of this study was to characterize peptidergic changes in the streptozotocin-induced diabetes mellitus rat model both quantitatively and qualitatively. Diabetes mellitus was induced by a single intraperitoneal injection of streptozotocin (65 mg/kg) and the levels of SP and CGRP were measured by means of radioimmunoassay (RIA) in a time-dependent manner. Peptide immunoreactivities were characterized by high pressure liquid chromatography (HPLC). The expression of both neuropeptides was examined 5 weeks after streptozotocin injection using in situ hybridization with 35S-labelled oligonucleotides. Saline-injected rats served as controls. SP was significantly decreased in the diabetic rat TG, i.e. , a 44.6% (+/-10.9) decrease after 1 week, 40.2% (+/-11.8) after 3 weeks and 72.3% (+/-14.6) after 5 weeks. CGRP was decreased only after 5 weeks (19.6% decrease +/-3.9), whereas at later stages, both peptide levels returned to normal values. HPLC revealed one major peak coeluting with the synthetic peptides. By using in situ hybridization, a significantly increased signal of both peptide-encoding mRNAs was found (43.8%), which seems to act to restore a diabetes-associated depletion of neuropeptides in the diabetic rat TG. The decreased SP- and CGRP levels in the diabetic rat TG reflect a diabetes-associated deficit which may be clinically relevant. Diabetes mellitus is associated with a variety of ocular complications, even corneal complications, including decreased corneal sensitivity, which in many ways resemble those after interruption of the normal trophic innervation of the eye. Our results point to reduced availability of neuropeptides for corneal innervation and may thus support the idea of a partial loss of trophic influences from the trigeminal nerve in diabetics.     

Ultrastructural identification of trigeminal nerve terminals in the pterygopalatine ganglion of rats: an anterograde tracing and immunohistochemical study

Trigeminal nerve terminals in the rat pterygopalatine ganglion (PPG) were ultrastructurally identified using anterograde tracing with Phaseolus vulgaris-leucoagglutinin (PHA-L). Electron microscopic immunohistochemistry was used to demonstrate the presence of substance P (SP) and calcitonin gene-related peptide (CGRP) in nerve terminals of the PPG. Adjacent to the rostral part of the PPG an additional minor area was described. Perikarya in this minor rostral part were more spherical and had irregular outlines. Ultrastructurally, the glial enwrapment of the nerve terminals seemed to be more loosely arranged in comparison to that in the major rostral part of the PPG. With PHA-L, numerous labelled nerve fibres and terminals were found in all parts of the PPG. The ultrastructure of these terminals was uniform, many of them showing synaptic contacts. Numerous terminals in the PPG were SP-positive, whereas only a few were CGRP-positive. Fibres stained positive for both neuropeptides. The PPG is shown to be synaptically innervated by sensory fibres arising in the trigeminal ganglion, with the strong suggestion of SP and CGRP acting as neurotransmitters. A modulatory interaction between the autonomic and sensory system, resembling an axon reflex mechanism in the peripheral nervous system is endorsed.     

Secretin mRNA in the Subdivision of Primary Sensory Neurons in the Trigeminal Ganglion of Rats

The primary sensory neurons use glutamate as a major neurotransmitter. Several neuropeptides are also found in these neurons. In our laboratory we demonstrated secretin-like immunoreactivity in primary sensory neurons of several species including human, rat and cat. In the present experiment utilizing in situ hybridization, we have demonstrated for the first time that secretin is not only immunostained but is also expressed in the primary sensory neurons of the trigeminal ganglion of male rats. In intact rats, secretin mRNA was not observed; we had to use intracerebroventricular colchicine administration to induce the expression of secretin. Secretin was expressed in about 5% of the cells in all the three subdivisions of the trigeminal ganglion. The secretin-synthetizing cells were large and medium sized, and their mean diameter was about 50 μm. When we compared the percentage and the size of secretin to that of calcitonin gene-related peptide (CGRP), substance-P (SP) and vasoactive intestinal polypeptide (VIP) cells, it was found that CGRP, SP and VIP are present in about 15–20% of the cells and their mean diameter is about 20–25 μm. The morphometric data indicate that secretin is present in a subdivision of neurons that is different from the subdivision of the CGRP, SP and VIP cells. It is suggested that secretin may modulate the function of the primary neurotransmitter.     

The effect of treatment with BRX-220, a co-inducer of heat shock proteins, on sensory fibers of the rat following peripheral nerve injury

In this study, we examined the effect BRX-220, a co-inducer of heat shock proteins, in injury-induced peripheral neuropathy. Following sciatic nerve injury in adult rats and treatment with BRX-220, the following features of the sensory system were studied: (a) expression of calcitonin gene-related peptide (CGRP); (b) binding of isolectin B4 (IB4) in dorsal root ganglia (DRG) and spinal cord; (c) stimulation-evoked release of substance P (SP) in an in vitro spinal cord preparation and (d) nociceptive responses of partially denervated rats. BRX-220 partially reverses axotomy-induced changes in the sensory system. In vehicle-treated rats there is a decrease in IB4 binding and CGRP expression in injured neurones, while in BRX-220-treated rats these markers were better preserved. Thus, 7.0 +/- 0.6% of injured DRG neurones bound IB4 in vehicle-treated rats compared to 14.4 +/- 0.9% in BRX-220-treated animals. Similarly, 4.5 +/- 0.5% of DRG neurones expressed CGRP in the vehicle-treated group, whereas 9.0 +/- 0.3% were positive in the BRX-220-treated group. BRX-220 also partially restored SP release from spinal cord sections to electrical stimulation of primary sensory neurones. Behavioural tests carried out on partially denervated animals showed that BRX-220 treatment did not prevent the emergence of mechanical or thermal hyperalgesia. However, oral treatment for 4 weeks lead to reduced pain-related behaviour suggesting either slowly developing analgesic actions or enhancement of recovery processes. Thus, the morphological improvement seen in sensory neurone markers was accompanied by restored functional activity. Therefore, treatment with BRX-220 promotes restoration of morphological and functional properties in the sensory system following peripheral nerve injury.     

The difference of osteocalcin-immunoreactive neurons in the rat dorsal root and trigeminal ganglia: co-expression with nociceptive transducers and central projection

The co-expression of osteocalcin (OC) with the capsaicin receptor (VR1) and vanilloid receptor 1-like receptor (VRL-1) was examined in the dorsal root (DRG) and trigeminal ganglia (TG). Virtually all OC-immunoreactive (ir) DRG neurons were devoid of VR1- and VRL-1-immunoreactivity (ir). In the TG, 14.1% of OC-ir neurons were also immunoreactive for VR1. Only 1.7% of OC-ir TG neurons co-expressed VRL-1-ir. The distribution of OC-ir was also examined in the spinal cord and trigeminal sensory nuclei. In the spinal cord, the superficial laminae of the dorsal horn were devoid of OC-ir. The neuropil was weakly stained in other regions of the spinal horns. The medullary dorsal horn (MDH) contained numerous OC-ir varicose fibers in laminae I and II. These fibers were occasionally observed originating from the spinal trigeminal tract. The neuropil was weakly stained in deeper laminae of the MDH, and the rostral parts of the trigeminal sensory nuclei. The present study suggests that OC-ir TG nociceptors send their unmyelinated axons to the superficial laminae of the MDH.     

Endothelin immunoreactivity and mRNA expression in sensory and sympathetic neurones following selective denervation

The localization of endothelin (ET) in perivascular nerve varicosities supports pharmacological evidence that ET is a neurotransmitter in the autonomic nervous system. To examine the potential source of ET previously localized in cerebrovascular nerves, ganglia which send projections to these vessels were immunolabelled for ET and examined at the ultrastructural level. The trigeminal (TG) and superior cervical ganglia (SCG) were examined in control rats and following either sensory denervation or sympathectomy. In control TG, ET immunolabelling was detected throughout the cytoplasm of a subpopulation of neurones whereas in the SCG only the occasional ET-positive neurone was seen. Following sensory denervation with capsaicin, very few ET-immunoreactive nerve cell bodies or nerve fibres were detected in the TG compared with control ganglia, suggesting that ET is predominantly localized in primary afferent neurones, although some remaining myelinated nerve fibres stained positively. ET labelling of neurones in the SCG was unaffected by sensory denervation. Following selective damage to sympathetic nerves with 6-hydroxydopamine, there was a marked increase in intensity of ET-labelling of nerve fibres in the TG, probably due to increased availability of nerve growth factor for sensory nerves. There was no effect on ET immunoreactivity in the nerve cell bodies and nerve fibres within the SCG. However, in situ hybridization techniques demonstrated that 6-hydroxydopamine sympathectomy resulted in a marked increase in ET-1 mRNA expression in the SCG neurones. In conclusion, sensory nerves projecting from the TG are a more likely source of ET-positive perivascular nerves in cerebral arteries than sympathetic nerves from the SCG. Damaged sympathetic neurones markedly increase ET mRNA expression. In view of the neuroprotective properties of ET, this may represent a compensatory mechanism to promote repair.     

Nasal mucosa sensitization with toluene diisocyanate (TDI) increases preprotachykinin A (PPTA) and preproCGRP mRNAs in guinea pig trigeminal ganglion neurons.

Toluene diisocyanate (TDI) induces respiratory allergy in mammals. Using immunohistochemistry and in situ hybridization histochemistry, the present study examined effects of nasal mucosa sensitization by TDI on the immunoreactivity for substance P (SP) and calcitonin gene-related peptide (CGRP) and on the expression of their mRNAs in guinea pig trigeminal ganglion and their terminals. Single intranasal application of TDI (acute experiment) did not induce nasal allergy-like behaviours and failed to cause changes of SP and CGRP immunoreactivity and in the expression of preprotachykinin A (PPTA) mRNA and preproCGRP mRNA coding for SP and CGRP respectively in the trigeminal ganglion neurons. However, repeated application of TDI (chronic experiment) caused a dramatic increase of SP and CGRP immunoreactivity in peripheral neurites of sensory nerves in the nasal mucosa but a slight increase in the spinal trigeminal nucleus, a decrease of the same immunoreactivities in the cell bodies of the trigeminal ganglion neurons, and an increase of the expression of PPTA and preproCGRP mRNA in the same neurons. These findings suggest that chronic exposure of the nasal mucosa to TDI apparently causes enhancement of both the biosynthesis of SP and CGRP and their axonal transport in the trigeminal system.     

The central and peripheral ends of the substance P-containing sensory neurones in the rat trigeminal system

The distribution of substance P (SP) immunoreactivity in the spinal nucleus of the rat trigeminal nerve and in the skin of the lower lip was examined following (a) unilateral electrolytic lesions of the trigeminal ganglion, (b) trigeminal rhizotomy, and (c) unilateral interruption of the mental nerve, the sensory branch of the trigeminal nerve innervating the lower lip. A marked depletion of SP immunoreactivity in the ipsilateral trigeminal spinal nucleus followed lesions of the trigeminal ganglion or rhizotomy. The reticular formation ventral and medial to the spinal nucleus showed a small decrease in SP immunofluorescence on the operated side. Some loss of SP immunoreactivity was observed in the skin of the lower lip following ganglionectomy or rhizotomy. After sectioning the mental branch SP-immunofluorescent fibres of the skin of the lower lip disappear completely on the denervated side. It was concluded that some trigeminal ganglion neurones store, and might release, SP at their axon terminals in the medulla oblongata and at their sensory terminals in the skin.     

Distribution of a splice variant of choline acetyltransferase in the trigeminal ganglion and brainstem of the rat: comparison with calcitonin gene-related peptide and substance P

Rat trigeminal ganglion neurons have been shown to contain a splice variant of choline acetyltransferase (pChAT). Here we report the distribution pattern of pChAT-containing afferents from the trigeminal ganglion to the brainstem, compared with that of calcitonin gene-related peptide (CGRP) and substance P (SP), by use of the immunohistochemical techniques in the rat. Most of CGRP(+) SP(+) ganglion cells contain pChAT, whereas half of the pChAT(+) ganglion cells possess neither CGRP nor SP. In the brainstem, pChAT(+) nerve fibers are found exclusively in the trigeminal and solitary systems, although the distribution pattern differs from that of CGRP(+) or SP(+) fibers. First, the ventral portion of the principal sensory nucleus contains many pChAT(+) fibers, with few CGRP(+) or SP(+) fibers. Because this portion receives projections of nociceptive corneal afferents, a subpopulation of pChAT(+) CGRP(-) SP(-) primary afferents is most probably nonpeptidergic nociceptors innervating the cornea. Second, the superficial laminae of the medullary dorsal horn, the main target of nociceptive afferents, contain dense CGRP(+) and SP(+) fibers but sparse pChAT(+) fibers. Because pChAT occurs in most CGRP(+) SP(+) ganglion cells, such sparseness of pChAT(+) fibers implies poor transportation of pChAT to axon branchlets. Another important finding is that pChAT(+) axons are smooth and nonvaricose, whereas CGRP(+) or SP(+) fibers possess numerous varicosities. Our confocal microscopy suggests colocalization of these three markers in the same single axons in some brainstem regions. The difference in morphological appearance, nonvaricose or varicose, appears to reflect the difference in intraaxonal distribution between pChAT and CGRP or SP.     

Glial cell line-derived neurotrophic factor (GDNF) from adult rat tooth serves a distinct population of large-sized trigeminal neurons

Glial cell line-derived neurotrophic factor (GDNF) mediates trophic effects for specific classes of sensory neurons. The adult tooth pulp is a well-defined target of sensory trigeminal innervation. Here we investigated potential roles of GDNF in the regulation of adult trigeminal neurons and the dental pulp nerve supply of the rat maxillary first molar. Western blot analysis and radioactive 35S-UTP in situ hybridization revealed that GDNF in the dental pulp and its mRNAs were localized with Ngf in the coronal pulp periphery, in particular in the highly innervated subodontoblast layer. Retrograde neuronal transport of iodinated GDNF and Fluorogold (FG) from the dental pulp indicated that GDNF was transported in about one third of all the trigeminal dental neurons. Of the GDNF-labelled neurons, nearly all (97%) were large-sized (> or =35 microm in diameter). Analysis of FG-labelled neurons revealed that, of the trigeminal neurons supporting the adult dental pulp, approximately 20% were small-sized, lacked isolectin B4 binding and did not transport GDNF. Of the large-sized dental trigeminal neurons approximately 40% transported GDNF. About 90% of the GDNF-accumulating neurons were negative for the high-temperature nociceptive marker VRL-1. Our results show that a subclass of large adult trigeminal neurons are potentially dependent on dental pulp-derived GDNF while small dental trigeminal neurons seems not to require GDNF. This suggests that GDNF may function as a neurotrophic factor for subsets of nerves in the tooth, which apparently mediate mechanosensitive stimuli. As in dorsal root ganglia both small- and large-sized neurons are known to be GDNF-dependent; these data provide molecular evidence that the sensory supply in the adult tooth differs, in some aspects, from the cutaneous sensory system.     

Colocalization of CGRP with 5-HT1B/1D receptors and substance P in trigeminal ganglion neurons in rats

Vasodilatation in the dura mater has been implicated in migraine pathogenesis. Anti-migraine triptan drugs block vasodilatation by binding to 5-HT1B/1D receptors localized on the peripheral sensory terminals and dural blood vessel smooth muscles. Previous studies suggest that calcitonin gene-related peptide (CGRP) released from Adelta-fibres plays a more important role than substance P (SP) released from C-fibres in inducing dural vasodilatation and that one of the antimigraine mechanisms of triptan drugs is inhibiting CGRP release. In the present study, the relationship between CGRP and 5-HT1B/1D receptors, and between CGRP and SP in the trigeminal ganglion neurons in rats was examined by double immunohistochemical staining. CGRP, 5-HT1B, 5-HT1D and SP-positive trigeminal ganglion neurons were all predominantly small and medium-sized. In the trigeminal ganglia, approximately 50% of CGRP-positive neurons were 5-HT1B positive. Similarly, approximately 55% of CGRP-positive neurons were 5-HT1D immunoreactive. Approximately 50% of CGRP-positive neurons were SP-positive, while 93% of SP-positive neurons were CGRP-positive, suggesting that nearly all SP-positive neurons also contain CGRP. The fibre types of the 5-HT1B- and 5-HT1D-positive neurons were further investigated with an antibody against the A-fibre marker 200-kDa neurofilaments (NF200). Approximately 46% of the 5-HT1B-positive and 43% of the 5-HT1D-positive trigeminal ganglion neurons were also NF200 positive, indicating that many A-fibre trigeminal neurons express 5-HT1B or 5-HT1D receptors. These results support the hypothesis that one important action of antimigraine drugs is the inhibition of CGRP release and that Adelta-fibres may play an important role in migraine pathogenesis.     

5-HT(1B) and 5-HT(1D) receptors in the human trigeminal ganglion: co-localization with calcitonin gene-related peptide, substance P and nitric oxide synthase

5-Hydroxytryptamine (5-HT) is implicated in migraine and agonist directed against 5-HT(1B) and 5-HT(1D) receptors are commonly used as effective therapies. The antimigraine mechanisms involve the inhibition of intracranial sensory neuropeptide release. In order to determine which 5-HT(1) receptor subtypes are involved we have by immunocytochemistry examined the distribution of 5-HT(1B) and 5-HT(1D) receptors in the human trigeminal ganglia, and addressed which of them colocalize with calcitonin gene-related peptide (CGRP), substance P (SP) or nitric oxide synthase (NOS). We detected that 5-HT(1D) receptor immunoreactivity (i.r.) was predominantly expressed in medium-sized cells (86% of positive cells, 30-60 microm). About 9% of the 5-HT(1D) receptor i.r. cells were large in size (> 60 microm) and 5% were small in size (< 30 microm). In a similar pattern, 5-HT(1B) receptor i.r. was mainly expressed in medium-sized cells (81% in 30-60 microm, 15% in > 60 microm and 4% in < 30 microm). Double immunostaining was used to determine whether the 5-HT(1B) or 5-HT(1D) receptor immunoreactive cells co-localized with either CGRP, SP or NOS. Thus, 89% of the CGRP i.r. cells expressed 5-HT(1D) receptor i.r. and 65% of the CGRP positive cells were 5-HT(1B) receptor positive. Most of the 5-HT(1D) (95%) and the 5-HT(1B) (94%) receptor i.r. cells showed SP immunostaining and 83% of 5-HT(1D) receptor and 86% of 5-HT(1B) receptor i.r. cells contained NOS. In conclusion, both 5-HT(1B) and 5-HT(1D) receptors are expressed in the human trigeminal ganglion and they are mainly localized in medium-sized cells and they seem to colocalize with CGRP, SP and NOS.     

α-Eudesmol, a P/Q-type Ca channel blocker, inhibits neurogenic vasodilation and extravasation following electrical stimulation of trigeminal ganglion

In this study, we investigated the effect of α-eudesmol, which potently inhibits the presynaptic ω-agatoxin IVA-sensitive (P/Q-type) Ca²⁺ channel, on neurogenic inflammation following electrical stimulation of rat trigeminal ganglion. Treatment with α-eudesmol (0.1–1 mg/kg. i.v.) dose-dependently attenuated neurogenic vasodilation in facial skin monitored by a laser Doppler flowmetry. In addition, α-eudesmol (1 mg/kg. i.v.) significantly decreased dural plasma extravasation in analysis using Evans blue as a plasma marker. On the other hand, α-eudesmol (1 mg/kg, i.v.) did not affect mean arterial blood pressure in rats. The calcitonin gene-related peptide (CGRP) and substance P (SP) released from activated sensory nerves have recently been suggested to be associated with the neurogenic inflammation. In this study, we also showed that α-eudesmol (0.45–45 μM) concentration-dependently inhibits the depolarization-evoked CGRP and SP release from sensory nerve terminals in spinal cord slices. These results indicate that the anti-neurogenic inflammation action of α-eudesmol, which does not affect the cardiovascular system, may be due to its presynaptic inhibition of the neuropeptide release from perivascular trigeminal terminals. We also suggest that the ω-agatoxin IVA-sensitive Ca²⁺ channel blocker, α-eudesmol, may become useful for the treatment of the neurogenic inflammation in the trigemino-vascular system such as migraine.     

The effect of sympathectomy on calcitonin gene-related peptide levels in the rat trigeminovascular system

The effect of sympathectomy on the calcitonin gene-related peptide (CGRP) level in the rat primary trigeminal sensory neurone was investigated. Six weeks after bilateral removal of the superior cervical ganglion there was a 70% rise in the CGRP content of the iris and the pial arteries, a 34% rise in the concentration in the trigeminal ganglion but no change in the brainstem. The CGRP rise in both end organs suggests that this phenomenon may be common to all peripheral organs receiving combined sensory and sympathetic innervations. The lack of any rise in the brainstem CGRP content raises the possibility that this process spares central terminations. In contrast, the level of neuropeptide Y, a peptide mainly contained in sympathetic terminals, fell to 35% of control values in the iris and pial arteries whilst the trigeminal ganglion and brainstem concentrations remained unchanged. The possible relevance of these observations to the clinical syndrome of postsympathectomy pain (sympathalgia) is discussed. There are similarities between the delayed onset of the human pain state and the delayed rise in sensory peptides after sympathectomy.     

Trigeminal inputs to reticulospinal neurones in lampreys are mediated by excitatory and inhibitory amino acids

Reticulospinal (RS) neurones integrate sensory inputs from several modalities to generate appropriate motor commands for maintaining body orientation and initiation of locomotion in lampreys. As in other vertebrates, trigeminal afferents convey sensory inputs from the head region. The in vitro brainstem/spinal cord preparation of the lamprey was used for characterizing trigeminal inputs to RS neurones as well as the transmitter systems involved. The trigeminal nerve on each side was electrically stimulated and synaptic responses, which consisted of mixed excitation and inhibition, were recorded intracellularly in the middle and posterior rhombencephalic reticular nuclei. The EPSPs were mediated by activation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors. An increase in the late phase of the excitatory response occurred when Mg²⁺ ions were removed from the Ringer's solution. This effect was antagonized by 2-amino-5-phosphonopentanoate (2-AP5) or reversed by restoring Mg²⁺ ions to the perfusate suggesting the activation of N-methyl-d-aspartate (NMDA) receptors. IPSPs were mediated by glycine. These findings are similar to those reported for other types of sensory inputs conveyed to RS neurones, where excitatory and inhibitory amino acid transmission is also involved.